Easy and Affordable: A New Method for the Studying of Bacterial Biofilm Formation.

BACKGROUND: Bacterial biofilm formation (BBF) proves itself to be in the spotlight of microbiology research due to the wide variety of infections that it can be associated with, the involvement in food spoilage, industrial biofouling and perhaps sewage treatment. However, BBF remains difficult to study due to the lack of standardization of the existing methods and the expensive equipment needed. We aim to describe a new inexpensive and easy to reproduce protocol for a 3D-printed microfluidic device that can be used to study BBF in a dynamic manner. METHODS: We used the SolidWorks 3D CAD Software (EducationEdition 2019-2020, Dassault Systèmes, Vélizy-Villacoublay, France) to design the device and the Creality3D Ender 5 printer (Shenzhen Creality 3D Technology Co., Ltd., Shenzhen, China) for its manufacture. We cultivated strains of Enterococcus faecalis, Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa. For the biofilm evaluation we used optical coherence tomography (OCT), scanning electron microscopy (SEM), Fourier Transform Infrared (FTIR) spectroscopy and crystal violet staining technique. RESULTS: Based on the analysis, Enterococcus faecalis seems to produce more biofilm in the first hours while Pseudomonas aeruginosa started to take the lead on biofilm production after 24 h. CONCLUSIONS: With an estimated cost around €0.1285 for one microfluidic device, a relatively inexpensive and easy alternative for the study of BBF was developed.

3D Printed Microfluidic Bioreactors Used for the Preferential Growth of Bacterial Biofilms through Dielectrophoresis.

A realistic modelling of the way biofilms form and evolve in time requests a dynamic approach. In this study, the proposed route uses continuous-flow bioreactors under controlled flow rates and temperature in the culture medium containing bacteria or fungi. 3D printed, Polylactic acid (PLA), flow-based bioreactors with integrated copper electrodes were used to investigate the effect of dielectrophoresis on the formation and growth of Staphylococcus aureus ATCC 25923, Enterococcus faecalis ATCC 29212, Pseudomonas aeruginosa ATCC 27853, and Klebsiella pneumoniae ATCC 13883 biofilms. Bacterial suspensions of 1McF turbidity have been prepared and circulated through the bioreactors. At the same time, a 30 V potential difference was applied on the system. The effect of the non-uniform electric field induced upon the bacterial cells was determined using quantitative methods, such as an adjusted microtiter plate technique, as well as spectral domain optical coherence tomography (SD-OCT) images. The morphology and the surface quality of the biofilms were investigated using Scanning Electron Microscopy (SEM) images. The results show that the different bacterial cells present a positive dielectrophoretic behaviour, with the preferential formation of biofilms in the high field gradient region.

Multilayered Porous Titanium-Based 3rd Generation Biomaterial Designed for Endosseous Implants.

This work proposes a novel complex multi-layered material consisting of porous titanium as a substrate and a complex coating consisting of a chitosan film engulfing microsphere loaded with growth factors such as BMP2 (bone morphogenic protein 2) and IGF1 (insulin-like growth factor-1). The microspheres were obtained through deposition of dual layers of calcium cross linked pectin-chitosan/pectin polyelectrolyte onto a BSA (bovine serum albumin) gel core. The multilayer was conceived to behave like a 3rd generation biomaterial, by slow delivery of viable growth factors around implants, and to assist the healing of implantation wound and the development of new vital bone. The biologic effect of the delivery of growth factors was studied in vitro, on MSC-CD1 mesenchymal stem cells, and in vivo, on CD1 mice. Proliferation and differentiation of cells were accelerated by growth factors, especially IGF1 for proliferation and BMP2 for differentiation. In vivo tests analyzed histologically and by MicroCT show a more structured tissue around BMP2 samples. The present concept will give the best clinical results if both growth factors are delivered together by a coating film that contains a double population of microcarriers.